Herpes simplex virus (HSV-1) causes diseases in humans, which can result in oral, genital, or ocular sores, or in rare occasions, life-threatening encephalitis. HSV-1 has two distinct phases of its viral life cycle: lytic and latent. An HSV-1 immediate-early protein, infected cell protein 0 (ICP0), is a key determinant as to whether an infection will be lytic or latent. ICP0 is a 110-KDa protein that potently transactivates viral gene expression via its E3 ubiquitin (Ub) ligase activity. Typically, E3 Ub ligases are components of pathways that attach and polymerize Ub (a 76 amino acid protein) to target proteins, marking them for degradation by the proteasome. Available evidence suggests that ICP0 mediates the degradation of several cellular proteins required for its transactivating activity. Efforts to identify specific targets of ICP0-directed degradation by proteomic approaches have been challenging because of limitations in the purity, solubility, and amount of a given protein required for its detection. While several functions of ICP0 during viral infection have been discovered, the identities of cellular proteins marked for degradation by ICP0 are largely unknown. Until these novel targets have been identified, it is unclear the exact role ICP0?s E3 Ub ligase activity plays in its transactivating activity. The long-term goal of our studies is to understand at the molecular level how virus-host interactions regulate the HSV-1 life cycle. The objective of this application is to a use a unique screening procedure to identify cellular proteins whose degradation is directed ICP0, determining the role these ICP0 targets play in HSV-1 gene expression and lytic replication. Our central hypothesis is that ICP0 mediates the degradation of novel cellular proteins; we propose that the loss of these proteins promotes efficient viral gene expression and replication. Our approach is to identify functional targets of ICP0-mediated degradation using a cutting edge screen. Our rationale for carrying out these studies is that by understanding how ICP0 facilitates productive infection via ubiquitination, targets of ICP0 degradation could be used to develop new therapies to impair HSV replication and its associated diseases. For this purpose, the following specific aims are proposed: Aim #1: Identify targets of the HSV-1 E3 Ub ligase, ICP0; Aim #2: Determine the contribution ICP0 targets play in HSV- 1 gene expression and replication. From this proposal we expect to identify cellular proteins involved in the biology of ICP0 using an innovative approach, which is a significant contribution that can used to isolate and identify genes and pathways that play important roles involved proteolysis and viral pathogenesis.